Probing the coordination behavior of Hg 2+ , CH 3 Hg + , and Cd 2+ towards mixtures of two biological thiols by HPLC-ICP-AES Katie L. Pei a , Melani Sooriyaarachchi a , Darren A. Sherrell b , Graham N. George b , Jürgen Gailer a, ⁎ a Department of Chemistry, University of Calgary, 2500 University Drive N.W., Calgary, Alberta, Canada T2N 1N4 b Department of Geological Sciences, University of Saskatchewan, Saskatoon, Saskatchewan, Canada S7N 5E2 abstract article info Article history: Received 9 July 2010 Received in revised form 16 November 2010 Accepted 26 November 2010 Available online 7 December 2010 Keywords: Cadmium(II) Mercury(II) Methylmercury Glutathione Liquid chromatography ICP-AES Hepatocyte cytosol contains a multitude of proteins, but also comparatively high concentrations of L-glutathione (GSH, ~5.0 mM) and L-cysteine (Cys, ~ 0.5 mM). Since Hg 2+ , CH 3 Hg + and Cd 2+ have a high affinity for thiols, their coordination to these thiols is likely involved in their intracellular transport. The comparative coordination behavior of these metal species towards mixtures of Cys and GSH, however, has not been studied under near physiological conditions. To probe these toxicologically relevant interactions, each metal species was separately injected onto a C 18 -HPLC column (37 °C) that had been equilibrated with phosphate buffered saline (PBS) that contained 5.0 mM GSH (mobile phase) and detected with an inductively coupled plasma atomic emission spectrometer. The incremental increase of the Cys concentration in the mobile phase (in 0.5 or 1.0 mM steps) up to 10 mM followed by the chromatography of each metal species decreased the retention of Hg 2+ and CH 3 Hg + albeit in a different manner. This behavior was rationalized in terms of the replacement of hydrophobic GS-moieties coordinated to each mercurial by less hydrophobic Cys-moieties. In contrast, a Cd-peak eluted close to the void volume with all investigated mobile phases. Using X-ray absorption spectroscopy, the Cd-compound that eluted with a PBS-buffer that contained 5.0 mM GSH was structurally characterized as tetrahedral (GS) 4 Cd. Thus, the in vivo formation of (GS) 4 Cd must be considered and HPLC-ICP-AES is identified as a useful tool to probe dynamic bioinorganic processes which involve the interaction of a metal ion with multiple ligands under physiologically relevant conditions. © 2010 Elsevier Inc. All rights reserved. 1. Introduction Mercury (Hg) and cadmium (Cd) are chemical constituents of the Earth's crust which have been continually released into the environ- ment for eons by natural processes, such as volcanic activity and the chemical weathering of minerals [1]. As a result, these toxic metals have always been present in surface and ground waters, soils, and the atmosphere at background concentrations. The progressive industri- alization of the world over the last two centuries, however, has led to an increased mobilization of these and other toxic metals from the earth's crust into the global environment [2]. In fact, the anthropo- genic emissions of Hg and Cd are currently estimated to be of the same order of magnitude as their natural emissions [1]. Consequently, certain human populations are exposed to higher levels of mercuric mercury (Hg 2+ ), methyl mercury (CH 3 Hg + ), and cadmium (Cd 2+ ) today via the ingestion of contaminated food, drinking water and/or the inhalation of polluted air than ever before [1,3–5]. 1 Due to the adverse health effects that are associated with the exposure of humans to low levels of toxic metals [1,6,7], the chronic exposure of the general population to mercurials 2 and Cd 2+ is increasingly recognized as an important public health issue [8–13]. The exposure of mammals to Hg 2+ , for instance, can lead to nephrotoxicity [8,14] and is possibly implicated in the etiology of neurological diseases, such as Parkinson's Disease [15,16]. Human exposure to CH 3 Hg + , on the other hand, can lead to neurotoxicity not only in adults [17,18], but also in the fetus, as CH 3 Hg + has been demonstrated to cross the placental barrier of pregnant women [19]. In contrast, Cd 2+ is classified as a human carcinogen [7,20,21] and the chronic exposure of humans to this metal can result in hepatoxicity [22] and/or nephrotoxicity [23,24]. Importantly, Cd 2+ has also been demonstrated to act as an endocrine disruptor after the administra- tion of mammals with extremely small doses [7]. Despite extensive research, however, the biomolecular basis for the chronic toxicity of Hg 2+ , CH 3 Hg + , and Cd 2+ in mammals is still not completely understood [1,5,14,25]. Once absorbed into the systemic blood circulation of a mammalian organism, toxic metals are distributed to various internal organs Journal of Inorganic Biochemistry 105 (2011) 375–381 ⁎ Corresponding author. Fax: + 1 403 289 9488. E-mail address: jgailer@ucalgary.ca (J. Gailer). 1 Throughout this manuscript, the use of Hg 2+ , CH 3 Hg + , and Cd 2+ implies a generic meaning and does not necessarily indicate that these species are present as cations. 2 Throughout the manuscript the term mercurials refers to Hg 2+ and CH 3 Hg + . 0162-0134/$ – see front matter © 2010 Elsevier Inc. All rights reserved. doi:10.1016/j.jinorgbio.2010.11.019 Contents lists available at ScienceDirect Journal of Inorganic Biochemistry journal homepage: www.elsevier.com/locate/jinorgbio